Hi
I'm wondering if anyone has cultured mixed glia on chamber slides. I'm using Nunc Tissue-Tek II 8 well or 4 well chamber slides, and whilst the cultures are doing well in Nunc flasks, there are barely any cells in the chambers.
Any help or advice would be appreciated!

-rachneuroimm-

try to coat with polylysine for attaching of the cells

are the bootoms of those chambers not made of glass? I think that might be the problem

-ggrrrmmmlllbbb-

I am also trying to grow them in T-75 flasks. They have been in culture 18 days and still only seeing maybe 2-3% confluency. The rest are floating in suspension and dying off or dead. I have never had issues with a cell line this hard before. The protocol I was using said they should attach after 5-7 days. I've tried non-coated, lysine coated, collagin coated and lamminin. The one that had the best results were the non-coated & lysine coated. But even w the coated flask, the improvement was negligible. Please let me know too if you find something that works! good luck!

-cmccray-

ggrrrmmmlllbbb on Tue Dec 21 15:00:26 2010 said:

try to coat with polylysine for attaching of the cells

are the bootoms of those chambers not made of glass? I think that might be the problem

Hi, Thankyou very much for your reply! I've only just got it as I thought this would email me if anyone responded :/
I did end up trying PLL coating and it worked great.
Thanks!

-rachneuroimm-

cmccray on Sat Feb 12 04:56:06 2011 said:

I am also trying to grow them in T-75 flasks. They have been in culture 18 days and still only seeing maybe 2-3% confluency. The rest are floating in suspension and dying off or dead. I have never had issues with a cell line this hard before. The protocol I was using said they should attach after 5-7 days. I've tried non-coated, lysine coated, collagin coated and lamminin. The one that had the best results were the non-coated & lysine coated. But even w the coated flask, the improvement was negligible. Please let me know too if you find something that works! good luck!

Have you had any more luck with this? My cultures are directed at growing OPCs but I'm using primary cells isolated from neonatal mouse brains and growing them in SATO media. What cell line are you using and what media? If there's astrocytes involved, they usually grow really well and you would expect the flask to be near enough covered with them by this point. If it's just OPCs or something else there may be much less and you might need to alter your media?